一种与细胞周期素A1互作的新基因的克隆和表达

目的：了解与细胞周期素Al互作的一个EST的组成结构和蛋白质细胞定位。方法：通过酵母双杂交试验获得了一个与细胞周期素Al互作的新EST，进一步获得了全长基因focal，其读码框长度为666bp，编码221个氨基酸。将该基因克隆进杆状病毒AcNPV及真核表达质粒pcDNA3中，分别进行了表达和定位。结果：获得了表达该基因的重组杆状病毒和融合了EGFP的真核表达质粒。蛋白印迹试验证明该基因在昆虫Tn5细胞中得到了表达；并发现该基因的产物在COS-7细胞中定向于细胞核内。结论：该蛋白与cyclin A1（核蛋白）存在互作的可能性。此结果为深入研究focal基因的功能打下了基础。

Molecular cloning and expression of a novel cyclin A1 interacting gene foca1

Objective:To understand the structure and location of an EST protein interacting with cyclin A1.Methods: By yeast triple-hybrid test,a new EST(foca1) was acquired,which might be a candidate interacting with cyclin A1.The foca1,a span gene,was obtained in further by RACE,with a length of ORF was 666 base pairs,encoding 221 amino acids.This gene was cloned into baculovirus AcNPV and pcDNA3 plasmid fusing to EGFP and then was expressed and located,respectively. Results: The recombinant baculovirus and plasmid were obtained.It demonstrated that the gene was expressed in insect Tn5 cells by Western-blot.The gene products was located in the nucleus of COS-7 cells tranfected with the recombinant plasmid pcDNA3-egfp-foca1. Conclusion: It indicated that foca1 had the interacting probability with Cyclin A1 which also was a nuclear protein.Further studies will reveal the function of the foca1 gene.